Aberrant regulation of cytokine signaling results in a wide variety of hyper-inflammatory, autoimmune and immune-deficiency pathological conditions. Cytokines are a large and diverse group of molecules which mediate interactions between cells, ultimately regulating the wide variety of cells of the immune repertoire. Cytokine signaling mediates numerous facets of normal immune system physiology, including development, response, activation, maintenance, memory and apoptosis (Roitt et al. (Eds.), Immunology, Fifth Edition, Mosby International Publishers [1998]).
Tumor Necrosis Factor
Tumor necrosis factor-α (commonly written as TNF, but also written as tumor necrosis factor or TNFα) is a multifunctional cytokine mediating pleiotropic biological functions in both health and disease states. TNF is secreted primarily by monocytes and macrophages, but can also be secreted by other cell types. The list of processes regulated by TNF is extensive, and includes inflammation, immunoregulation, cyytotoxicity and antiviral effects (See e.g., Vilcek et al, J. Biol. Chem., 266:7313–7316 [1991]). TNF plays an integral role in destroying tumors, mediating responses to tissue injury, and protecting hosts from infections by various microorganisms (Vassali, Ann. Rev. Immunol., 10:411–452 [1992]). TNF also induces the transcriptional activation of numerous genes, including NF-κB and AP-1, with the consequent expression of pro-inflammatory and immunoregulatory genes (Rothe et al., Cell 83:1243–124 [1995]; Varfolomeev et al., J. Exp. Med., 183:1271–1275 [1996]; Chinnaiyan et al., J. Biol. Chem., 271:4961–4965 [1996]; Hsu et al., Immunity 4:387–396 [1996]; and Hsu et al., Cell 84:299–308 [1996]). TNF-mediated NF-κB activation is also an important negative feedback mechanism regulating apoptosis (Beg and Baltimore, Science 274:782–784 [1996]; Van Antwerp et al., Science 274:787–789 [1996]; and Wang et al., Science 274:784–787 [1996]).
TNF in Disease
TNF has also been implicated in the pathogenesis of a variety of diseases and disorders. It is theorized that these pathologies result from the aberrant regulation of TNF activity, in which the pathologies manifest as a result of excessive or insufficient TNF activity. Among the activities for which TNF is most noted are its pro-inflammatory actions, sometimes termed the “acute phase immune response.” Unfortunately, if not properly regulated, these proinflammatory responses can result in tissue injury and chronic inflammatory diseases, such as rheumatoid arthritis, inflammatory bowel disease, septic shock, cachexia, autoirumune disorders, graft-versus-host disease and insulin resistance (Piguet et al., J. Exp. Med., 166:1280 [1987]; Pujol-Borrell et al., Nature 326:304–306 [1987]; Tracey et al., Nature 330:662–664 [1987]; Oliff, Cell 54:141–142 [1988]; Vilcek et al., J. Biol. Chem., 266:7313–7316 [1991]; and Eigler et al., Immunol. Today 18:487–92 [1997]). Excessive TNF activity results in the detrimental effects of an exaggerated immune response demonstrated in some of these diseases, exemplified by overstimulation of interleukin-6 and granulocyte/macrophage-colony stimulating factor (GM-CSF) secretion, enhanced cytotoxicity of polymorphonuclear neutrophils, prolonged expression of cellular adhesion molecules, induction of procoagulant activity on vascular endothelial cells, increased adherence of neutrophils and lymphocytes, and stimulation of the release of platelet activating factor from macrophages, neutrophils and vascular endothelial cells (Vassali, Ann. Rev. Immunol., 10:411–452 [1992]; Vilcek et al., J. Biol. Chem., 266:7313–7316 [1992]; and Barbara et al., Immunol. and Cell Biol., 74:434–443 [1996]).
Recent evidence also implicates TNF activity in the pathogenesis of many infections. TNF is thought to play a central role in the pathophysiological consequences of Gram-negative sepsis and endotoxic shock, including fever, malaise, anorexia, and cachexia (Beutler et al., Nature 316:552–554 [1985]; Bauss et al, Infect. Immun., 55:1622–1625 [1987]; Tracey et al., Nature 330:662–664 [1987]; and Vassali, Ann. Rev. Immunol., 10:411–452 [1992]). Because TNF can mimic many of the biological effects of endotoxin, it is theorized that TNF is a central mediator responsible for the clinical manifestations of endotoxin-related and other critical illnesses (Waage et al., Lancet 1:355–357 [1987]; Cerami et al., Immunol. Today 9:28 [1988]; Mitchie et al., N. Eng. J. Med., 318:1481–1486 [1988]; Revhaug et al., Arch. Surg., 123:162–170 [1988]; and Michie et al., Ann. Surg., 209:19–24 [1989]).
Tumor Necrosis Factor Receptor
The numerous biological effects of TNF are now known to be mediated by two transmembrane receptors, the 55 kilodalton Type I receptor (also written as “CD120a,” and referred to herein as “TNFR1”) and the 75 kilodalton Type II receptor (also written as “CD120b,” and referred to herein as TNFR2). Although both TNFR1 and TNFR2 demonstrate strong affinity for TNFα, these two receptors demonstrate no apparent homology in their cytoplasmic (i.e., intracellular) domains. This fact is consistent with the observation that these two receptors transduce different signals to the nucleus via distinct signaling intermediates (Lewis et al., Proc. Natl. Acad. Sci. USA 88:2830–2834 [1991]; Tartaglia and Goeddel, Immunol. Today 13:151–153 [1992]; and Barbara et al., Imunol. Cell Biol., 74:434–443 [1996]).
Soluble TNF inhibitors have been identified in normal human urine, as well as in sera and other body fluids of patients with infectious, neoplastic and immunologic disorders. This observation ultimately led to the revelation that these soluble TNF inhibitors were actually the extracellular domains of TNF receptors derived by proteolytic cleavage of the transmembrane forms (Engelmann et al., J. Biol. Chem., 264:11974–11980 [1989]; Olsson et al., Eur. J. Haematol., 42:270–275 [1989]; Seckinger et al., J Biol. Chem., 264:11966–11973 [1989]; Engelmann et al., J. Biol. Chem., 265:1531–1536 [1990]; and Aderka et al., J. Exp. Med., 175:323–329 [1992]). In the case of the TNFR1, this proteolytic activity results in the cleavage and shedding of the extracellular N-terminal domain (also called the ectodomain). These free, soluble TNFR1 ectodomains (“sTNFR1s”) have an affinity for TNF that is similar to that of intact membrane receptors. Due to this affinity, the free receptors are able to bind and sequester TNF, thereby inhibiting the biological action of TNF. Furthermore, the generation of sTNFR1 is also likely to suppress TNF signaling by reducing the number of functional TNF receptors acting at the cell membrane. The sTNFR1 ectodomains are also theorized to serve a more complex buffering function in the regulation of TNF activity (Aderka et al., J. Exp. Med., 175:323–329 [1992]; and Werb and Yan, Science 282:1279–1280 [1998]). The complexity of TNF signaling is fuirther illustrated by the observation that many of the stimuli that result in TNF release also result in the release of the soluble TNF receptor, suggesting that these soluble TNF inhibitors may serve as part of a regulated feedback mechanism to control TNF activity (Adreke et al., J. Exp. Med., 175:323–329 [1992]; and Porteu and Nathan, J. Exp. Med., 172:599–607 [1990]).
The importance of TNFR1 in the regulation of TNF activity in host defense, immunoregulation and development has been fuirther demonstrated in studies utilizing TNFR1 knockout mice. Mice deficient in TNFR1 show a variety of phenotypes, including phenotypes which mimic human immune disorders (Pfeffer et al., Cell 73:457–467 [1993]; Rothe, Nature 364:798–802 [1993]; Le Hir et al., J. Exp. Med., 183:2367–2372 [1996]; Matsumoto et al., Science 271:1289–1291[1996]; Mori et al. J. Immunol. 157:3178–3182 [1996]; Speiser et al., J. Immunol., 158:5185–5190 [1997]; Tkachuk et al., J. Exp. Med., 187:469–477 [1998]; and Kagi et al., J. Immunol., 162:4598–4605 [1999]).
The key role of TNFR1 shedding in the regulation of TNF bioactivity is highlighted by the association of germline mutations in TNFR1 extracellular domains with impaired TNFR1 shedding and autoinflammatory disease characterized by autosomal dominant periodic fever syndromes (McDermott et al., Cell 97:133–144 [1999]).
Other Mediators of Acute Phase Response
In addition to TNF, other cytokines have been implicated in the induction of the pro-inflammatory response (i.e., the acute phase immune response). These cytokines which demonstrate overlapping activities with TNF include the interleukins (e.g., IL-1 and IL-6) (Suffredini et al., J. Clin. Immunol., 19:203–214 [1999]).
IL-1 (consisting of both α and β forms) is an important proinflammatory cytokine which regulates the expression of a wide variety of target genes and proteins in nearly every cell type (Dinarello, Blood 77:1627–1652 [1991]; and Dinarello, The Cytokine Handbook (ed. Angus W. Thomson), 3rd edition, Academic Press, San Diego, p. 35–72 [1998]). The spectrum of IL-1-mediated biologic effects includes inflammatory, metabolic, physiologic, hematopoietic, and immunologic functions. IL-1 is thought to play a role in the pathogenesis of several disease states, including septic shock, rheumatoid arthritis, inflammatory bowel disease, myelogenous leukemia, diabetes mellitus, and atherosclerosis (Dinarello et al., N. Engl. J. Med., 328:106–113 [1993]).
IL-6 is also a multifunctional cytokine with pleiotropic pro-inflammatory effects (DiCosmo, et al., J. Clin. Invest., 94:2028–2035 [1994]; and Kishimoto et al., Blood 86:1243–1254 [1995]). For example, IL-6 plays an important role in regulating B cell immunoglobulin production, T-cell activation, growth and differentiation, hematopoiesis, hepatic acute phase reactions and osteoclast development (Hirano, The Cytokine Handbook (ed. Angus W. Thomson), 3rd edition, Academic Press, San Diego, p. 197–228 [1998]). Dysregulated production of IL-6 may contribute to the pathogenesis of a variety of inflammatory, neoplastic and autoimmune disorders, such as plasma cell neoplasia and Castleman's disease (Yoshizaki, et al., Blood 74:1360–1367 [1989]; and Hirano, Int. J. Cell Cloning 9:166–184 [1991]).
The signal transduction pathways utilized by TNF, IL-1 and IL-6 also show shared signaling intermediates. For example, both TNF and IL-1 can activate both NF-κB and AP-1, which are important pro-inflammatory transcription factors (Ashkenazi et al., Science 281:1305–1308 [1998]; and Dinarello, The Cytokine Handbook (ed. Angus W. Thomson), 3rd edition, Academic Press, San Diego, p. 35–72 [1998]). Similarly, IL-6 signaling uses components of the JAK-STAT pathway, which has also been reported to be induced by TNF (Guo et al., J. Immunol., 160:2742–2750 [1998]; and Hirano, The Cytokine Handbook (ed. Angus W. Thomson), 3rd edition, Academic Press, San Diego, p. 197–228 [1998]).
The cognate receptors for the IL-1 and IL-6 cytokines are known. There are two IL-1 receptor forms, type I and type II. There is a single IL-6 receptor, consisting of gp80 alpha chain and gp130 beta chain subunits, where ligand binding is mediated by the alpha subunit. The IL-1 and IL-6 receptors are also present as soluble forms analogous to the soluble form of TNFR1. Furthermore, it has been suggested that these receptors play a role in the regulation of IL-1 and IL-6 activity and pro-inflammatory response (Dower et al., J. Immunol., 142:4314 [1989]; Novick et al., J. Exp. Med., 170:1409 [1989]; Eastgate et al., FEBS Lett., 260:213 [1990]; Giri et al., J. Biol. Chem., 265:17416 [1990]; Symons et al., Cytokine 2:190 [1990]; Symons et al., FEBS Lett., 272:133 [1990]; Symons et al., J. Exp. Med., 174:1251–1254 [1991]; Mullberg et al., Biochem. Biophys. Res. Commun., 189:794 [1992]; Mullberg et al., Eur. J. Immunol., 23:473 [1993]; Svenson et al., Cytokine 5:427 [1993]; and Arend et al., J. Immunol., 153:4766–4774 [1994]).
Analogy between regulation of TNF and other cytokines is further illustrated by studies utilizing peptide-hydroxamate metalloprotease inhibitors. Specifically, the protease inhibitors TAPI (TNF-α protease inhibitor) and RU36156 have been reported to inhibit the proteolytic cleavage and shedding of both TNFR1 and IL-6R (Mullberg et al., J. Immunol., 155:5198–5205 [1995]; and Gallea-Robache et al., Cytokine 9:340–346 [1997]).
As discussed above, in view of the importance of TNF, IL-1 and IL-6 in both health and disease states, there exists a need for methods and compositions for the regulation of TNF, IL-1 and IL-6 cytokine activity. These methods and compositions will find use as therapeutic agents for the treatment of disease states.